Changes in the magnitude of the shear stress acting on the walls of blood vessels cause alterations in endothelial permeability, endothelial cell morphology and gene expression. Increased shear stress results in increased arachidonic acid metabolism with the primary metabolic being prostacyclin. PGI2, in human umbilical vein endothelial cells, HUVECs. Prostacyclin causes vasodilation, modulates vascular permeability, inhibits platelet aggregation and suppresses immune responses. The rate limiting step of arachidonic acid metabolism to PGI2 is catalyzed by Prostaglandin H synthase (PGHS). The increase in PGI2 production caused by an increase in shear stress along with the dependence of PGI2 on PGHS, suggests that PGHS gene expression may be regulated by shear stress. Two different genes designated PGHS1 and PGHS2 have previously been isolated and sequenced. Specific Aims 1 and 2 of this proposal is to characterize in HUVECs the effect of shear stress on the levels of PGHS1 and PGHS2 mRNA. Confluent monolayers of cells will be exposed to shear stresses of 4, 15 and 25 dyn/cm2 for up to 24 hr. The message levels will be quantified using quantitative RT-PCR. Specific Aim 2 is to identify the PGHS2 shear stress regulatory element. Fusion genes comprised of the human growth hormone and regions of the PGHS2 promoter will be used to locate the regulatory element. Specific Aim 4 is to determine the mechanisms involved in the regulation of the genes by shear stress. The effect of inhibiting second messengers like protein kinase C and calcium on the regulation of the genes by shear stress will be determined. This research will increase our understanding of how mechanical forces effect the function of endothelial cells.